Fig. 4: C/EBPα a is required to maintain but not select AT2 fate.

a Bioinformatic screen of 1678 transcriptional or epigenetic regulators, of which 67 are enriched along AT2 lineage. 32 of these have known embryonic lethal or lung defects upon knockout in mice and can be stratified into four temporal categories: DP, DP-nAT2, nAT2-mAT2, and constitutive. The observed plasticity window is depicted in red. b Dot plot of Sox9 (DP restrictive; yellow), Cebpa and Nupr1 (nAT2-mAT2 restrictive; pink), Etv5 and Elf5 (constitutive; gray) temporal expression (plasticity window in red). c E16.5 and E17.5 lungs immunostained for PDPN (white), C/EBPα (green), ECAD (red), and DAPI (blue). C/EBPα⁺ cells are marked (arrowheads) within the proximal (P) and distal (D) regions. Bars, 100 µm. d Alveolar regions of PN0 control Nkx2.1^Cre; Rosa26^mTmG; Cebpa^wt/fl (left) and Nkx2.1^Cre; Rosa26^mTmG; Cebpa^fl/fl (right) lungs immunostained for DP lineage reporter (GFP) and MUC1 (white). Note GFP⁺ AT2s and AT1s (flat and MUC1⁻, labeled “1”) form even after Cebpa deletion. Bars, 50 µm. e Alveolar regions of adult control LyzM^Cre; Rosa26^mTmG; Cebpa^wt/fl (left) and LyzM^Cre; Rosa26^mTmG; Cebpa^fl/fl (right) lungs immunostained for MUC1 (white) and CD74 (blue). AT2-lineage derived GFP⁺ cells differentiate into AT1s (asterisk) upon Cebpa deletion. Bars, 100 µm. f Quantification of (e) showing percent GFP⁺ cells that differentiate into AT1s upon Cebpa deletion (n represents number of GFP⁺ cells sampled for the Cebpa^wt/fl and Cebpa^fl/fl conditions in experimental triplicate). ****p =  1.4 × 10⁻¹⁴ (Student’s two-sided t test, data as mean ± SD). All experiments were repeated at least three times. Source data are provided as a Source Data file.